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Phorbol ester activation of chloride current in guinea‐pig ventricular myocytes
Author(s) -
Shuba Lesya M.,
Asai Tatsuya,
MCDonald Terence F.
Publication year - 1996
Publication title -
british journal of pharmacology
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 2.432
H-Index - 211
eISSN - 1476-5381
pISSN - 0007-1188
DOI - 10.1111/j.1476-5381.1996.tb15298.x
Subject(s) - protein kinase c , depolarization , voltage clamp , patch clamp , phorbol , reversal potential , biophysics , myocyte , chemistry , membrane potential , guinea pig , medicine , endocrinology , electrophysiology , biochemistry , biology , kinase
1 Although earlier studies with phorbol esters indicate that protein kinase C (PKC) may be an important regulator of Cl − current ( I Cl ) in cardiac cells, there is a need for additional quantitative data and investigation of conflicting findings. Our objectives were to measure the magnitude, time course, and concentration‐dependence of I Cl activated in guinea‐pig ventricular myocytes by phorbol 12‐myristate 13‐acetate (PMA), evaluate its PKC dependence, and examine its modification by external and internal ions. 2 The whole‐cell patch clamp technique was used to apply short depolarizing and hyperpolarizing pulses to myocytes superfused with Na +‐ , K + ‐, Ca 2+ ‐free solution (36°C) and dialysed with Cs + solution. Stimulation of membrane currents by PMA (threshold ≤1nM, EC 50 14nM, maximal 40% increase with ≥100 nM) plateaued within 6–10 min. 3 PMA‐activated current was time‐independent, and suppressed by 1 mM 9‐anthracenecarboxylic acid (9‐AC). Its reversal potential ( E rev ) was sensitive to changes in the Cl − gradient, and outward rectification of the current‐voltage ( I‐V ) relationship was more pronounced with 30 mM than 140 mM Cl − dialysate. 4 The relative permeability of PMA‐activated channels estimated from E rev measurements was I − >Cl − » aspartate. Channel activation was independent of external Na + . 5 PMA failed to activate I Cl in myocytes pretreated with 1‐(5‐isoquinolinesulphonyl)‐2‐methylpiperazine (H‐7) or dialysed with pCa 10.5 solution. Lack of response to 4α‐phorbol 12, 13‐didecanoate (αPDD) was a further indication of mediation by PKC. 6 I Cl induced by 2 μ m forskolin was far larger than that induced by PMA, suggesting that endogenous protein kinase A is a much stronger Cl − channel activator than endogenous PKC in these myocytes. 7 The macroscopic properties of PMA‐induced I Cl appear to be indistinguishable from those of PKA‐activated I Cl . We discount stimulation of PKA by PMA as an explanation, and conclude that endogenous PKC may activate PKA‐regulated Cl − channels in these myocytes.